CRMB, Recombinant, Variola (Cytokine Response-Modifying Protein B) (BSA Free)

Variola virus is the causative agent of human smallpox. Like other poxviruses, it encodes a variety of molecules that shield virus infected cells from immune clearance. The cytokine response modifiers CRMB, C, D, and E, which are differentially expressed among the poxviruses, function as decoy TNF receptors and block the proinflammtatory and antiviral effects of TNF (1, 2). Of the CRM proteins, Variola virus encodes only CRMB, secreted from virus infected cells as a 90kD disulfide linked dimer (3). The N-terminal 112 amino acid (aa) region of CRMB mediates binding to human, mouse, and rat TNF as well as human lymphotoxin-alpha, and neutralizes the cytolytic effects of TNF (3, 4). The C-terminal 155 aa region of CRMB, known as a SECRET domain (smallpox virus-encoded chemokine receptor), binds the chemokines CCL25, CCL28, CXCL12b, CXCL13, and CXCL14, which are involved in the antiviral immune response (4). Functionally, the SECRET domain interferes with the in vitro migration of T cells in response to CCL25 (4). A SECRET domain is also present in CRMD but not in CRMC or CRME. Variola virus CRMB shares 84%-92% aa sequence identity with camelpox virus, cowpox virus, and monkeypox virus CRMB, but only 21% with vaccinia virus CRMB (which lacks a SECRET domain). The TNF binding domain of CRMB shares 30% and 42% aa sequence identity with comparable regions of human TNF R1 and R2, respectively. Source: DNA sequence encoding the mature Variola CRMB sequence (Ala 23-Leu 349, Accession # P34015) was fused with a 6X histidine tag at the C-terminus and expressed in a mouse myeloma cell line, NS0. , Definition: Measured by its ability to inhibit the TNF-alpha mediated cytoxicity in the mouse L-929 cell line, in the presence of the metabolic inhibitor actinomycin D. Applications: Suitable for use in Western Blot. Other applications not tested. Recommended Dilution: Western Blot: Predicted 36.5kD protein migrates as an ~ 55-60kD protein in SDS-PAGE under reducing conditions., Optimal dilutions to be determined by the researcher. Storage and Stability: Lyophilized powder may be stored at 4°C for short-term only. Reconstitute to nominal volume by adding sterile PBS and store at -20°C. Reconstituted product is stable for 12 months at -20°C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.